Dialogue between Professor Jerry Nadler and Yumi Imai

Their discussion is based on the recent review article “Interaction between cytokines and inflammatory cells in islet dysfunction, insulin resistance and vascular disease” by Drs Imai, Dobrian, Nadler and others, published in Diabetes, Obesity and Metabolism 15 (Suppl. 3): 117-119, 2013.

Their discussion is based on the recent review article “Interaction between cytokines and inflammatory cells in islet dysfunction, insulin resistance and vascular disease” by Drs Imai, Dobrian, Nadler and others, published in Diabetes, Obesity and Metabolism 15 (Suppl. 3): 117-119, 2013.

Yumi Imai: We would like to discuss the therapeutic potential of 12 lipoxygenase (12LO) inhibitors, for both type 1 and type 2 diabetes. To start with, what is a 12LO pathway?

Jerry Nadler: 12LO is an enzyme expressed in many key tissues relevant to diabetes in humans and animal models of diabetes. It converts fatty acids, particularly arachidonic acids, into signaling lipid metabolites. 12-hydroxyeicosatetraenoic acid (12 HETE) is one of the very damaging and relatively stable metabolites. 12LO in humans, which is also called Alox 12 enzyme, was originally named platelet 12 lipoxygenase because it is highly expressed in platelets and to some extent in megakaryocytes. However, we were the first group to show that an Alox12 enzyme is the major lipoxygenase in human pancreatic b-cells and highly expressed in visceral adipose tissue, especially in diabetic humans. It is also expressed in inflammatory cells including macrophages, in the brain, and the colon. Dr. Imai, maybe you can share your experience in the islet, as we are trying to identify where in the islet the 12LO is expressed, because it may not only be in the b-cell.

Yumi Imai: Interestingly, 12LO seems to be extremely high in cells that produce pancreatic polypeptides in individuals at early stages of type 2 diabetes, and those atantibody-positive and prediabetic stages of type 1 diabetes. It is very intriguing that 12LO seems to be high in islets during a certain stage of the diabetes, for both type 1 and type 2 diabetes.

Considering the nature of 12LO, I think 12LO inhibition falls into the category of anti-inflammatory therapy. There are some other therapies, like IL-1b and NFkB targeted pathways, being studied for diabetes. Where does 12-LO inhibition fall compared with, or in relation to, other anti-inflammatory therapies?

Jerry Nadler: The 12LO pathway can be activated through IL-1b, NFkB, and others. But it appears to be unique in that it amplifies many inflammatory pathways. When 12LO gets activated, it could activate NADPH oxidase which is a very potent enzyme pathway leading to oxidative stress. The 12LO pathway also leads to endoplasmic reticulum (ER) stress, which is thought to be a very common and potentially important pathway leading to b-cell dysfunction or maybe b-cell death. It also activates certain inflammatory MAP kinases and other inflammatory cytokines. Therefore, by targeting 12LO, you could get multiple benefits by blocking cytokine production, MAP kinases, ER stress, and even oxidative stress.

Yumi Imai: What is the evidence supporting the role of 12LO specifically in type 1 diabetes based on animal and human studies?

Jerry Nadler: One of the challenges for the study of 12LO has been the lack of selective and potent inhibitors. We provided the first evidence that 12LO pathway is involved in type 1 diabetes using low-dose streptozotocin to simulate type 1 diabetes in a global 12LO knockout mouse. The animal was highly protected from inflammatory injury independent of nitric oxide. More recently, we crossed the 12LO knockout mouse on to the NOD spontaneous diabetes background. The results were very striking showing almost complete protection and prevention of type 1 diabetes in the NOD mouse when you deleted 12LO. The newest work ongoing is using mouse with floxed 12LO so that we can make targeted knockouts. We have current fundings from the Juvenile Diabetes Foundation, making targeted knockout mice by either deleting 12LO in the macrophage or in the pancreas in the NOD background. In addition to in vivo studies, we have studied human histology provided by nPOD showing staining of the 12LO in prediabetic and early type 1 diabetic donors. For in vitro studies, islets isolated from the 12LO knockout mice are protected from the cytokines that are linked to development of type 1 diabetes. We feel there is a good amount of evidence supporting the role of 12LO in type 1 diabetes. We are now looking to develop new inhibitors, especially small molecules that we can use in the animal model, and see if we can prevent diabetes or prevent the full development of diabetes.

Yumi Imai: How about type 2 diabetic islets? Recently, evidence is emerging that supports the role of inflammation in islet demise in type 2 diabetes. Would you think that the islet inflammation in type 2 is similar to type 1, and can they be targeted by 12LO inhibition?

Jerry Nadler: As you rightly suggest, more and more data suggests that inflammation in islets of type 2 diabetes is playing a role in b-cell loss. So, we are looking in samples of islets from type 2 diabetic humans. We are also studying an animal model, the db/db mouse, looking at 12LO expression at different time points in the islet and other tissues. The data is very suggestive that the Alox12 form of 12LO is highly upregulated in islet at the time when the b-cells are starting to get killed. I know Dr Imai, that you are doing work in the human tissues.

Yumi Imai: Yes, I think the db/db data and human data seem to agree very well in a sense that 12LO activation is seen at the critical stage during the development of type 2 diabetes. For humans, it is impossible to obtain islets as type 2 diabetes develops in real time. However, we are very fortunate to get human islets from large number of donors affected by type 2 diabetes. When we evaluated their function as glucose stimulated insulin secretion, those who have mild impairment in first phase insulin secretion are the ones with high 12LO expression. Actually, islets that are highly damaged and have lost response to glucose do not show elevated 12LO expression, but show upregulation of TNF a or MCP-1. So, 12LO may be a good target to preserve the islet function in those at the critical time before islets lose all their function. For type 2 diabetes, in addition to islet dysfunction, insulin resistance is another key pathology, and therapeutics that addresses both insulin resistance and islet dysfunction would be ideal. From that prospect, I think 12-LO has a lot of promising data.

Jerry Nadler: I agree. We have focused on adipose tissue as there is a clear link between visceral obesity, development of insulin resistance, and development of diabetes and cardiovascular disease. In collaboration with Dr Anca Dobrian at Eastern Virginia Medical School, we have obtained exciting, interesting data, suggesting that the 12LO pathway is upregulated particularly in the visceral adipose tissue, not subcutaneous adipose tissue in obese humans. Additional data showed that visceral adipose tissuie from rodent models over-express 12LO in diet-induced obesity and insulin resistance.

Yumi Imai: What happens if you delete it?

Jerry Nadler: When global 12-LO knockout mouse is put on a high-fat diet, our group and one other group in San Diego (they did it with glucose clamp; we did it with IV glucose tolerance test and the insulin tolerance test), both found the improvement in insulin resistance. Interestingly enough, the mice got just as obese as wild-type. They gained the same amount of weight, but they didn’t show inflammation in the adipose tissue with that. Then, we were very fortunate to cross the 12LO floxed mouse with the AP2 cre deleter mouse. We basically could delete 12LO in adipose tissue but also maybe some macrophage. We found that the mouse is protected from insulin resistance during a high-fat “Western” type diet. In addition, just by deleting 12LO in the adipose tissue, we protected against inflammation in islets, which indicates some crosstalk between the two tissues. We are moving on to use other deleter mice where we can selectively delete an adipocyte directly or in islets in collaboration with Dr Raghu Mirmira’s group at Indiana University.

Yumi Imai: How about the liver?

Jerry Nadler: I think that the other key tissue we haven’t really studied is the liver. As we know, during the metabolic syndrome, you get the fatty liver that is a major contributor to insulin resistance. A study group in Spain found that the 12LO global knockout mouse on a high-fat diet does not develop fatty liver.

Yumi Imai: Recently betatrophin has been reported as an interesting peptide produced by the liver and adipose tissue that regulates b-cell mass. It will be interesting to determine whether the expressions of betatrophin is altered by12 LO deletion.

To end our discussion, is 12LO inhibition expected to reduce cardiovascular complications? As you know, cardiovascular benefits will increase the value of any new diabetic therapeutics.

Jerry Nadler: That is a very good point. That was actually looked at earlier by researchers who specialize in atherosclerosis. When the 12LO gene was eliminated from Apoe null mouse, LDL receptor null mouse, or combined hyperlipidemia mouse models, they all showed clear protection from atherosclerosis. Bone marrow transplantation studies suggested the macrophage is involved, but also non-macrophage tissues seem to contribute to the protection against atherosclerosis. To conclude, ample evidence now exists for the involvement of 12LO activation in atherosclerosis as well as insulin resistance and b-cell dysfunction. Once a good inhibitor becomes available, it might have some benefits for both the metabolic and cardiovascular components of diabetes pathology.